Учебники / Genetics and Auditory Disorders Keats 2002

2B.J.B. Keats and C.I. Berlin

Awareness that inheritance is important in hearing impairment can be traced to the sixteenth century (see Nance and Pandya, Chapter 5, for a summary of these early studies), and more recently, Konigsmark (1969), Konigsmark and Gorlin (1976), Fraser (1976), and Gorlin et al. (1995) provided comprehensive reviews of hereditary hearing impairment and associated abnormalities emphasizing the extensive clinical and genetic variability. Estimates of the number of genes causing inherited hearing impairment (Stevenson and Cheeseman 1956; Chung and Brown 1970) varied depending on the size of the survey and the genetic heterogeneity of the particular population studied (Morton 1991).

One purpose of this volume in the Springer Handbook of Auditory Research series is to inform the reader about the many clinical forms of genetic hearing loss, the mutations that are responsible, and the functions of the proteins that are encoded by the mutant genes. However, the editors and authors view with equal importance the basic principles of genetics, the methods and techniques used in human molecular genetic studies, and the examples of how scientific results can be applied in patient care. Chapters 2 (Avraham and Hasson) and 3 (Giersch and Morton) describe the structure of genes and chromosomes, while Chapters 3 and 4 (Mueller, Van Camp, and Lench) explain the methodologies and the genomic tools used to find the chromosomal locations and identify disease genes. In Chapter 5, Nance and Pandya discuss the application of the laws of population genetics and models of genetic epidemiology to hearing loss, while Chapter 8 (Steel, Erven, and Kiernan) clarifies the relevance of mouse models to advancing understanding of hearing loss in humans. The clinical and genetic heterogeneity of genetic hearing loss are delineated in Chapters 6 (Griffith and Friedman) and 7 (Fischel-Ghodsian), with Chapter 6 providing an in-depth coverage of autosomal and X-linked forms of hearing loss and Chapter 7 concentrating on hearing loss due to mitochondrial DNA mutations. Finally, Chapter 9 (Arnos and Oerlich) illustrates the role of the genetic counselor in communicating knowledge about hearing loss to the patient, thus translating the findings of medical science into clinical care.

An overview of the basic principles of genetics is provided in this chapter, together with citations to other chapters that expand on the topic. In addition, this chapter ends with an extensive glossary of genetic terms with which some readers may be unfamiliar. More comprehensive coverage of human genetics may be found in textbooks such as Nussbaum et al. (2001) and Gelehrter et al. (1998). But before getting to the genetics of hearing loss, the phenotype requires both a definition and a discussion.

1.1 Defining and Quantifying the Hearing

Loss Phenotype

The findings that result from a clinical examination are an individual’s phenotype. They may include biochemical, physiological, and morphological characteristics. This phenotype can result from genetic factors, environmental factors, or a combination of both. The genetic factors are the individual’s genotype. Hearing loss may be just one of the features of the phenotype, in which case the disorder is known as a syndrome; if no other clinical anomalies are apparent, the hearing impairment is called nonsyndromic. Gorlin et al. (1995) describe 30 syndromes in which hearing loss is associated with abnormalities of the external ear, 40 with the eye, 87 with the musculoskeletal system, 23 with the kidney, 56 with the skin, 63 with the central nervous system, 51 with endocrine and metabolic conditions, 12 with chromosomal anomalies, 8 with oral and dental problems, and 35 with miscellaneous disorders, as well as 22 forms of nonsyndromic hearing impairment. The phenotypes and genotypes of many of these syndromes, as well as those of nonsyndromic hearing loss, are described in Chapters 6 and 7. The reader is able to keep updated on new hearing loss genes by accessing the Hereditary Hearing Loss Homepage (URL: http://dnalab-www.uia.ac.be/dnalab/hhh/).

The hearing loss phenotype has been most commonly quantified by obtaining a pure tone air conduction audiogram. The work that led to the development of the equipment necessary to make these measurements was done over 70 years ago. Georg von Békésy had just reported his theory on the traveling wave in the cochlea and its role in frequency analysis in 1929 (see Békésy 1960), when Crowe et al. (1934) published their paper showing that high-frequency hearing loss was associated with loss of hair cells at the base of the cochlea. The convergence of these independent lines of investigation led to considerable excitement in the field. Up to that time, hearing loss had been estimated with mechanical devices such as a monochord (a bow-activated, carefully calibrated string), whistle (Bunch 1943), and, of course, a tuning fork. What was unmistakable, however, was that Crowe et al. (1934) had used what was then a revolutionary instrument to quantify hearing loss. Their tool was the Western Electric 1-A audiometer, a battery driven device with an oscillator that produced pure tones up to 16,384 Hertz (Hz), and an interrupter, attenuator, and black bakelite earphones, whose output verification was difficult to quantify at the time. However, the voltage to the terminals of the phones was quite constant, and hearing losses were expressed both in decibels (dB) of attenuation and dB relative to what was thought to be normal sensitivity at octave intervals from 64 to 16,384 Hz. Thus, Crowe et al. (1934) were able to quantify hearing loss in dB, although at the time they were less interested in clinical issues than in harvesting bones with quantifiable hearing loss. It was some time before norms were developed, but this early 1930s electronic audiome-

4B.J.B. Keats and C.I. Berlin

ter predated audiology as it is practiced today and was the cutting edge of auditory assessment in humans.

The pure tone air conduction audiogram is at best only a rudimentary tool for measuring hearing loss. Today’s physiologically based audiological battery should include tympanometry, middle ear muscle reflexes, otoacoustic emissions, speech audiometry, and Auditory Brainstem Response/ Electrocochleography (ABR/EcochG). Auditory Brainstem Response and Electrocochleography are computer-controlled averaged evoked responses that delineate synchronous neural discharges, usually following a brief auditory pulse. They correlate extremely well with hearing, although they are not themselves hearing tests (Hood and Berlin 1986). Understanding why these additional tests are essential for fully accurate phenotyping requires review of the following basic principles of auditory physiology.

The five major electroacoustic events in the cochlea are:

1.The endocochlear potential

2.The cochlear microphonic

3.The compound action potential

4.The summating potential

5.Otoacoustic emissions

Selective abnormalities in one or a number of these elements can lead to 120 different permutations of these five events, although current level of knowledge can rule out a number of impossible sequences. For example, absence of endocochlear potential leads to an absence of all the other electrical events, including the otoacoustic emissions. It is therefore impossible, as far as we can tell today, to have an absent endocochlear potential and recordable otoacoustic emissions, or any of the above events. Removing such impossible sequences reduces the number of possible permutations to about 70.

Nowadays, parts of the organ of Corti can be assessed separately. For example, normal otoacoustic emissions indicate that the endocochlear potential, the middle ear, and the (primarily) outer hair cells are working well. ABR and EcochG provide information about the synchrony and integrity of the inner hair cells and eighth nerve, as well as confirming neural synchrony after the discharge of the primary neurons. Thus, these tests and their successors should be used not just in identifying the presence or absence of tumors, but also in assessing the integrity of the auditory nervous system and helping develop methods to categorize various genetic hearing losses audiologically. The same pure-tone audiogram might be observed in one patient with a genetically based loss of outer hair cells, and another with normal outer hair cells, but disturbed neural synchrony (Berlin et al. 1993; Starr et al. 1996). However, etiology and genotypes are quite different, and classifying both patients according to their pure-tone audiograms alone would not be adequate for genetic analysis. A more precise categorization of the hearing loss can be made by testing otoacoustic emissions,

6B.J.B. Keats and C.I. Berlin

providing a total of 46 chromosomes (22 autosomal pairs plus XX or XY) in the nucleus of a cell. If both parents transmit an X chromosome, the offspring is female (XX); if the father transmits a Y chromosome, then the offspring is male (XY). The distribution of family members with hearing loss may be consistent with autosomal or X-linked inheritance, which may be further categorized as dominant or recessive. The symbol assigned to a nonsyndromic form of hearing loss reflects the mode of inheritance. DFNA* and DFNB* (*the asterisk represents a number that is assigned when the gene for the hearing loss is localized to a chromosome) indicate autosomal dominant and autosomal recessive, respectively, while DFN* means X- linked. The number does not reflect the chromosomal location; DFNB7, for example, simply means that the gene for this form of autosomal recessive hearing loss was the seventh to be localized (the DFNB7 gene is on chromosome 9). If the pattern of inheritance is consistent with maternal transmission, the location of the gene may be the mitochondrial genome (Fischel-Ghodsian, Chapter 7). For hearing loss, there is general agreement that the pattern of inheritance is autosomal recessive in about 77% of cases, autosomal dominant in about 22%, and X-linked or mitochondrial in the remainder (Gorlin et al. 1995).

An individual has pairs of chromosomes and, therefore, pairs of genes and pairs of genetic markers (fragments of DNA that have unique chromosomal locations). The chromosomal location of a gene or genetic marker is its locus. The different forms of genes and genetic markers are called alleles; thus, at each locus an individual has two alleles, and this pair of alleles is the genotype. The genotype of the individual is homozygous if the two alleles are the same, and heterozygous if they are different.

1.2.1 Autosomal Dominant Inheritance

The pattern of inheritance is probably autosomal dominant if individuals in each generation are affected; both males and females are equally likely to be affected. Affected individuals are usually heterozygotes with one normal and one deleterious copy of the gene for the disorder, and each offspring of an affected individual has a 50% chance of inheriting the deleterious allele. However, knowing the genotype may not be adequate in predicting the phenotype. There may be variable expression among affected individuals, and some who must have the deleterious allele (because, for example, they have an affected parent and an affected child) may show no phenotypic signs.

Penetrance is defined as the probability of expressing features of the phenotype given the presence of a particular genotype. For a dominant disorder, if 70% of individuals who have the deleterious allele show some phenotypic expression of the disorder, then the penetrance value is 0.7. Penetrance may also be age dependent, such as with adult-onset hearing loss.

In this case, the probability that a person who has the deleterious allele will

have hearing loss increases with age. Thus, penetrance may be close to zero at 10 years of age, 0.3 at 20 years of age, 0.7 at 30 years of age, and 1.0 at 50 years of age. The degree of penetrance needs to be taken into account when calculating recurrence risks for individuals who have relatives with an autosomal dominant disease. Incomplete penetrance and/or variable expression among family members suggests that other genes and environmental factors are contributing to the phenotype.

An offspring with an autosomal dominant disorder may sometimes have two unaffected parents. As discussed above, this situation is observed with incomplete penetrance (a value less than 1.0). If, in addition, the affected offspring has no affected relatives, then the possibility of a new mutation must be considered. For example, more than 80% of cases of achondroplasia (short-limbed dwarfism) are new mutations in germ cells (sperm or ova). Occasionally, unaffected parents have more than one offspring with a dominant disorder. A likely explanation in this situation is germline mosaicism, which occurs if the mutation arose during the embryonic life of one of the parents in a cell destined to be a germ cell. As a result, some (or possibly all) of the germ cells carry the mutation.

1.2.2 Autosomal Recessive Inheritance

The mode of inheritance of a disorder is autosomal recessive if the abnormal phenotype is expressed only in individuals who have two copies of the deleterious allele. Heterozygotes are not affected and are often called carriers. Assuming complete penetrance, the probability that a child of carrier parents will be affected is one quarter; the probability that an unaffected sibling is a carrier is two thirds (Fig. 1.1). If a hearing impaired child of normal hearing parents has an affected sibling or the parents are related, then the etiology is most likely to be a deleterious gene with the mode of inheritance being autosomal recessive. Alternatively, if a known environmental insult such as an infectious agent or a drug is documented, then the presence of a deleterious gene is less likely. However, the possibility of recessive inheritance must always be considered. Note that family history is quite likely to be negative with a recessive disorder because the abnormal allele may be passed from one (unaffected) carrier to the next for many generations before a couple, who by chance both carry the same deleterious allele, has an affected child. Recessive inheritance probably explains most cases of profound sensorineural hearing loss when family history is negative, and there is no known environmental factor that could be responsible.

It is not uncommon for deaf individuals to marry one another. However, the underlying causes of their deafness may be different, even though both may have recessive deafness. If both are homozygous for the same deleterious allele, then all their children will also be homozygous for this allele.

However, if deleterious alleles for different genes are responsible for the

8B.J.B. Keats and C.I. Berlin

FIGURE 1.1. Recessive deafness. The probabilities (in parentheses) of each of the possible offspring phenotypes and genotypes are shown for parents who are both carriers.

deafness in the spouses, then all their offspring will probably be heterozygous for two deafness genes, and not homozygous for either of them. Thus, deaf spouses, who may each have many deaf relatives, can have offspring with normal hearing.

1.2.3 X-linked Inheritance

Males have both an X and Y chromosome; females have two X chomosomes. Thus, a necessary condition for X-linked inheritance is no father- to-son transmission because a son receives his X chromosome from his mother. If a disorder has an X-linked dominant pattern of inheritance with complete penetrance, then all daughters of affected fathers are affected, and each child of an affected mother has a 50% chance of being affected. In the case of an X-linked recessive trait, the majority of affected individuals are male because they have only one X chromosome. Females have two X chromosomes and may be carriers, but are unlikely to be affected. Sons of a carrier mother each have a 50% chance of inheriting the abnormal allele, while daughters have a 50% chance of being carriers. Note that, although females have two X chromosomes in a cell, in general one of the two is randomly inactivated early in embryonic development. Thus, females are mosaic, with each cell having one or the other X chromosome active.

1.2.4 Mitochondrial Inheritance

Mitochondria are small organelles that are located within the cytoplasm of a cell; they are independent of the nucleus and have their own DNA

(mtDNA). A process called oxidative phosphorylation (OXPHOS) takes place in the mitochondria and is responsible for energy (ATP) production in the cell. Each mitochondrion has multiple copies of mtDNA and each cell contains several hundred mitochondria, and thus many copies of mtDNA. The mutation rate is ten times higher in mtDNA than in nuclear DNA, and cells may contain both mutant copies and normal copies, a condition known as heteroplasmy. If the normal copies can successfully provide the energy requirements of the cell, then normal function will be retained.

However, different cell types have differing energy requirements. The organs with the most demand for energy are the skeletal muscle, heart, eye, ear, and brain. Thus, the typical symptoms found in mitochondrial disorders are muscle weakness, nervous system disorders, visual problems, hearing loss, and dementia.

The mtDNA molecule is circular and consists of about 16,000 base pairs. Mitochondria are maternally inherited, so the expected family history for a mitochondrial disorder is that all children of an affected mother are affected, and children of an affected father are never affected. However, because of heteroplasmy, the mother and siblings of an affected individual may show only minor symptoms, or none at all. Mitochondrial disorders that involve hearing loss are discussed by Fischel-Ghodsian (Chapter 7).

2. The Genetic Material

Each chromosome consists of a single molecule of deoxyribonucleic acid (DNA), which has a double-helical structure. DNA is composed of a sugarphosphate backbone and four bases: adenine (A) and guanine (G) are purines, and thymine (T) and cytosine (C) are pyrimidines. The double helix is formed through the pairing of A with T, and C with G, and the bases are held together by hydrogen bonds. Thus, knowing the sequence of bases on one DNA strand automatically gives the sequence on the other strand. This precise pairing means that DNA can replicate by separation of the two strands, followed by each strand serving as a template for a new complementary strand.

The set of 24 chromosomes has three billion base pairs, of which 1% to 2% make up the estimated 35,000 human genes. Genes consist of both coding sequences (exons) and intervening sequences (introns), and it is the exons that encode the amino acid sequence of the protein product. The connecting link between the gene (DNA) and the protein is messenger ribonucleic acid (mRNA). The sequence of bases in the exons is called the complementary DNA (cDNA) sequence because it can be synthesized using the mRNA as a template. The three major differences between DNA and RNA are: (1) DNA is double-stranded, RNA is single-stranded; (2) the